We have designed and implemented metabolite profiling studies, in which we incubate the compounds of interest with human hepatocytes and identify metabolites via high-resolution mass spectrometry (QTOF 5600). This cooperation project with ABSciex and Bristol Myers Squibb has produced an outcome of six published papers describing the metabolism of different new synthetic cannabinoids (XLR-11, AKB-48, RCS-4, RCS-8, STS-135, PB-22/5F-PB-22, and several others) and providing mass spectra for the scientific community to identify use of these drugs. Currently, studies on AH-7921, a new synthetic opioid, and three more synthetic cannabinoids AB-FUBINACA, AB-PINACA/5F-AB-PINACA are performed. Since spring 2014, we also established the analytical procedures for the metabolism identification studies on a second instrument, the Thermo QExactive, and are currently processing four synthetic cathinones alpha-PVT, alpha-PBP, 4-MeO-alpha-PVP and PV8. Complementing the human hepatocyte studies we designed and implemented human liver microsome experiments to assess the metabolic stability (half-life) of new designer drugs, which will provide a first estimate of how quickly the compound is metabolized. In the spring of 2014, we instituted a cooperation with Molecular Discovery, who provide software for in silico prediction of metabolites by simulating 3D docking of molecules in the enzymes catalytic cavity and consider reactive sites in the molecule. Evaluating the softwares potential will help advancing future metabolism studies. We also developed several analytical assays to identify new designer drugs in human specimens: Qualitative assays with library search were validated for 29 synthetic cannabinoids on LC-MS/MS (QTrap 5500) and for an even wider range of synthetic cannabinoids with the new SWATH approach using the QTOF5600. Quantitative assays were validated for 53 synthetic cannabinoids using LC-MS/MS (QTrap 5500) and for 28 cathinones using Orbitrap technology (QExactive). The latter method was recently extended to include 33 cathinones, 3 amphetamines and 8 piperazines. A quantitative analytical method for MDPV and two metabolites was developed to determine MDPV pharmacokinetics in rats after multiple administration routes. Our goal is to continually monitor the appearance of novel psychoactive substances on the US market and worldwide markets and to perform metabolite identification studies on prevalent new compounds. The goal is to publish our findings and provide MS/MS spectra to the scientific community in a timely manner to facilitate development of analytical methods. We also plan to extend the scope of the human liver microsome experiments by performing experiments with individual CYP450 enzymes and elucidate pathways and predict interactions. 3. As part of the collaboration with Dr. Baumann we are developing a quantitative assay for the synthetic cannabinoid AM2201 and its metabolites, which will be used to determine pharmacokinetics of this compound in rats. 4. A long-term goal is to prepare and eventually perform a controlled administration study with a synthetic cannabinoid in humans. To do so, preclinical pharmacology and toxicity tests have to be run first to file for an Investigational New Drug Application, then a protocol can be submitted.